This project will investigate the mechanism of the active transport system, or pump, for sodium and potassium ions across animal cell membranes. Ion translocation is coupled to phosphorylation of sodium, potassium transport adenosine triphosphatase, (Na,K)ATPase, from ATP with release of ADP and inorganic phosphate. The phosphate group of the phosphoenzyme is a most sensitive and available reporter group of the reactive state of the enzyme. The project will investigate the effect of high concentrations of sodium ion, up to 3 molar, on the kinetics of formation and breakdown of the phosphoenzyme and on the equilibrium binding of nucleotides. This approach is based on the presupposition that release of sodium ion from the enzyme (to the solution in contact with the extracellular face of the membrane) can be revented in this way and that reactive states antecedent to that release can be isolated. The enzyme will be prepared from the outer medulla of mammalian kidney by the zonal method of Jorgenesen. Since vanadate (V) is a powerful allosteric inhibitor and since it is reduced to vanadyl ion (IV) intracellularly, the power of vanadyl ion to inhibit the enzyme will be investigated. The project will seek experiments to demonstrate functional interactions between alpha peptide chains of this enzyme. The project will compare active site phosphopeptides of this enzyme with those of gastric (H ion, K ion) transport ATPase.